1. Field of the Invention
The present invention relates generally to the field of molecular pathology. More specifically, the present invention relates to a method for creating frozen tissue arrays for subsequent assaying.
2. Description of the Related Art
The implementation of high-throughput genetic technologies, such as oligonucleotide microarrays, generates myriad points of data. The identified potential candidate genes need to be further characterized and selected using a large number of well-characterized tumors and stringent criteria. Tissue microarrays allow for such high-throughput expression profiling of tumor samples, additionally providing information at the microanatomical level.
Sections cut from tissue arrays allow parallel detection of DNA, e.g., by fluorescence in-situ hybridization (FISH), RNA, e.g., by mRNA in-situ hybridization (mRNA ISH) or protein, e.g., by immunohistochemistry (IHC) targets in each of the multiple specimens in the array. Each microarray block can be sectioned up to 200-300 times. Therefore, tens of thousands of tissue microarray sections can be obtained from a set of tissue specimens in one recipient block. This substantially facilitates molecular profiling of very large numbers of cancer tissues and allows the generation of large-scale correlative databases, including clinical information and molecular data (including images), while ensuring that the donor blocks from which the tissue cores are removed can continue to be utilized so that research materials are not destroyed.
In early 1998, Kononen et al. (1) described a tissue microarray “chip” that had been developed for high-throughput molecular profiling of tumor specimens. Tissue microarrays enable rapid in-situ analysis of up to 1000 tumors or other tissues in a single experiment. In the method of Kononen, original tissue sample sources are morphologically representative regions of regular formalin-fixed paraffin-embedded tumor blocks. Core tissue biopsies are taken from individual “donor” paraffin-embedded tumor blocks and precisely arrayed into a new “recipient” paraffin block using a custom built instrument. Thereafter, Bubendorf et al. published data of a survey of gene amplifications during prostate cancer progression by high-throughput fluorescence in-situ hybridization on tissue microarrays (2). The first hand-held paraffin tissue array apparatus was later marketed.
Tissue microarrays consisting of 0.6 mm biopsies of paraffin-embedded tissues have been used for various clinicopathological studies. This size is sufficient for assessing morphological features of the analyzed tissues on many samples. However, the size of the biopsy used in these arrays may not be representative of the whole tumor specimen because of tissue heterogeneity. Additionally, paraffin tissue arrays have distinct limitations in maintaining intact RNA transcription levels, as well as proteins and other molecules (i.e., lipids) due to the fixatives and chemical reagents required for the paraffin process. Thus, tissue microarray technology using paraffin-embedded tissues can reach its limits for the detection of RNA targets or certain proteins. The use of a frozen tissue array or a cryoarray strategy would overcome these limitations and would allow for the processing of multiple frozen tissue specimens and/or cell lines on a single tissue block.
Therefore, it would be beneficial to have an effective means and a system for creating tissue arrays that allow all molecules to be assayed at the expression level and simultaneously visualized at micro-anatomical levels. Specifically, the prior art is deficient in the lack of an effective method for creating a frozen tissue array for subsequent assaying. The present invention fulfills this long-standing need and desire in the art.